TY - JOUR
T1 - A multisubstrate reductase from Plantago major
T2 - structure-function in the short chain reductase superfamily
AU - Fellows, Rachel
AU - Russo, Christopher M.
AU - Silva, Catarina S.
AU - Lee, Soon Goo
AU - Jez, Joseph M.
AU - Chisholm, John D.
AU - Zubieta, Chloe
AU - Nanao, Max H.
N1 - Funding Information:
This work was supported by Action Thématique et Incitative sur Programme (ATIP)-Avenir and Grenoble Alliance for Cell and Structural Biology (ANR-10-LABX-49-01) (to C.Z. and C.S.S.).
Publisher Copyright:
© 2018, The Author(s).
PY - 2018/12/1
Y1 - 2018/12/1
N2 - The short chain dehydrogenase/reductase superfamily (SDR) is a large family of NAD(P)H-dependent enzymes found in all kingdoms of life. SDRs are particularly well-represented in plants, playing diverse roles in both primary and secondary metabolism. In addition, some plant SDRs are also able to catalyse a reductive cyclisation reaction critical for the biosynthesis of the iridoid backbone that contains a fused 5 and 6-membered ring scaffold. Mining the EST database of Plantago major, a medicinal plant that makes iridoids, we identified a putative 5β-progesterone reductase gene, PmMOR (P. major multisubstrate oxido-reductase), that is 60% identical to the iridoid synthase gene from Catharanthus roseus. The PmMOR protein was recombinantly expressed and its enzymatic activity assayed against three putative substrates, 8-oxogeranial, citral and progesterone. The enzyme demonstrated promiscuous enzymatic activity and was able to not only reduce progesterone and citral, but also to catalyse the reductive cyclisation of 8-oxogeranial. The crystal structures of PmMOR wild type and PmMOR mutants in complex with NADP+ or NAD+ and either 8-oxogeranial, citral or progesterone help to reveal the substrate specificity determinants and catalytic machinery of the protein. Site-directed mutagenesis studies were performed and provide a foundation for understanding the promiscuous activity of the enzyme.
AB - The short chain dehydrogenase/reductase superfamily (SDR) is a large family of NAD(P)H-dependent enzymes found in all kingdoms of life. SDRs are particularly well-represented in plants, playing diverse roles in both primary and secondary metabolism. In addition, some plant SDRs are also able to catalyse a reductive cyclisation reaction critical for the biosynthesis of the iridoid backbone that contains a fused 5 and 6-membered ring scaffold. Mining the EST database of Plantago major, a medicinal plant that makes iridoids, we identified a putative 5β-progesterone reductase gene, PmMOR (P. major multisubstrate oxido-reductase), that is 60% identical to the iridoid synthase gene from Catharanthus roseus. The PmMOR protein was recombinantly expressed and its enzymatic activity assayed against three putative substrates, 8-oxogeranial, citral and progesterone. The enzyme demonstrated promiscuous enzymatic activity and was able to not only reduce progesterone and citral, but also to catalyse the reductive cyclisation of 8-oxogeranial. The crystal structures of PmMOR wild type and PmMOR mutants in complex with NADP+ or NAD+ and either 8-oxogeranial, citral or progesterone help to reveal the substrate specificity determinants and catalytic machinery of the protein. Site-directed mutagenesis studies were performed and provide a foundation for understanding the promiscuous activity of the enzyme.
UR - http://www.scopus.com/inward/record.url?scp=85054463784&partnerID=8YFLogxK
U2 - 10.1038/s41598-018-32967-1
DO - 10.1038/s41598-018-32967-1
M3 - Article
C2 - 30287897
AN - SCOPUS:85054463784
SN - 2045-2322
VL - 8
JO - Scientific Reports
JF - Scientific Reports
IS - 1
M1 - 14796
ER -